[Evaluation of carbapenem inactivation method for the identification of carbapenemase-producing Enterobacteriaceae strains]. / Enterobacteriaceae suslarinin saptanmasinda karbapenem inaktivasyon yönteminin degerlendirilmesi.

The rapid and accurate identification of carbapenemases is of crucial importance in terms of infection control. Methods employed in the determination of carbapenemases should be constantly updated in the light of technical advances and newly emerging carbapenemase variants. The aim of this study was to evaluate the performance of the newly developed carbapenem inactivation method (CIM) for the identification of carbapenemases defined in the members of the family Enterobacteriaceae. Enterobacteriaceae isolates with resistance to at least one of the carbapenems (ertapenem, imipenem or meropenem) were included in the study. The study isolates were obtained from various clinical specimens between 2008-2014 and consisted of 56 Enterobacteriaceae strains (12 Escherichia coli, 32 Klebsiella spp., and 12 Enterobacter spp.) in which the presence of the 38 blaOXA-48, 8 blaVIM, 7 blaIMP, 1 blaNDM-1, 1 blaKPC-2 and 1 blaOXA-48+blaVIM genes had been previously determined using the polymerase chain reaction (PCR) and 78 in which no carbapenemase gene were detected. For the performance of the CIM, the test bacteria were suspended in sterile water and then a 10 µg meropenem disc was immersed in the suspension and incubated for 2 hours. This meropenem disc was then removed and subsequently placed on a Mueller-Hinton agar plate inoculated with E. coli ATCC 29522 and incubated at 35°C. The results were assessed after 6 hours and after overnight incubation. Development of an inhibition zone around the meropenem disk was interpreted as the absence of carbapenemase and the lack of an inhibition zone as the presence of carbapenemase. The results of the CIM were obtained after 8 hours. With the CIM, all isolates with previously determined carbapenemase genes were found to be positive and the isolates with no genes revealed to be negative. The sensitivity and specificity of CIM were estimated as 100%. The high sensitivity and specificity, ease of application and interpretation, rapid production of results, and no necessity for additional equipment or chemical substances, makes CIM a promising high throughput method to be used in routine microbiology laboratories. Since this method indicates only the presence of a carbapenemase in a clinical isolate, the type of the carbapenemase present should be further identified by the use of molecular techniques.

[Comparison of the modified Hodge test and the Carba NP test for detection of carbapenemases in Enterobacteriaceae isolates]. / Enterobacteriaceae izolatlarinda karbapenemazlarin saptanmasinda modifiye Hodge testi ve Carba NP testlerinin karsilastirilmasi.

A rapid, practical, and accurate identification of carbapenemase-producing Enterobacteriaceae isolates is crucial for the implementation of appropriate infection control measures and proper treatment of the infections. For this purpose, a large number of phenotypic test methods have been developed, although none has 100% sensitivity and specificity. Variations in sensitivity and specificity of these tests based on the type of beta-lactamase enzymes carried by that isolates might result in differences between regions and countries. The aim of this study was to compare the performances of widely used modified Hodge test (MHT) and Carbapenemase Nordmann-Poirel (Carba NP) test in the detection of carbapenemases in Enterobacteriaceae family members. A total of 65 Enterobacteriaceae isolates (43 bla(OXA-48), 10 bla(VIM), 9 bla(IMP), 1 bla(NDM-1), 1 bla(KPC-2) and 1 bla(OXA-48)+bla(VIM) carrying strains) that showed decreased sensitivity to at least one carbapenem (ertapenem, imipenem or meropenem), and carriage of carbapenemase gene confirmed by polymerase chain reaction (PCR), were included in the study. Seventy-eight isolates showing decreased susceptibility to carbapenems but lacking carbapenemase genes were used as controls. All isolates were identified by using conventional methods as well as automated BD Phoenix System (Becton Dickinson, USA). The antimicrobial susceptibility testing was performed using the same automated system, and was confirmed by disk diffusion method. Results were evaluated according to the CLSI criteria. MHT was performed in accordance with the CLSI guideline, and Carba NP test was carried out by a modified protocol. Instead of imipenem monohydrate, which was used in the original protocol, 6 mg/ml imipenem/cilastatin was used in the modified protocol. In the study, MHT identified 90.8% (59/65) of carbapenemase-producing isolates, while 93.9% (61/65) of the isolates were identified by Carba NP test. With MHT, four Klebsiella pneumoniae producing OXA-48, one Escherichia coli producing IMP, and one K.pneumoniae producing NDM-1, and with Carba NP test, one E.coli and one K.pneumoniae producing OXA-48, one E.coli producing IMP, and one Enterobacter cloacae producing VIM could not de detected. Three OXA-48-producing isolates (two K.pneumoniae and one E.coli) yielded late and weak positive results with Carba NP test. MHT had false positivity for 31 isolates, while Carba NP test showed no false positivity. In comparison of the sensitivity and specificity of the two tests, sensitivities were found to be similar although the Carba NP has a slightly higher sensitivity than the MHT (93.9% versus 90.8%, respectively; p= 0.754), Carba NP was found more specific (100% versus 60.3%, respectively; p< 0.0001). With Carba NP test, 26% of the isolates (n= 16) were positive within 15 minutes, and 85% (n= 52) were positive within the first hour. It was concluded that, Carba NP test showed high sensitivity and specificity than the MHT and the results can be obtained more rapidly for the presence of carbapenemases in Enterobacteriaceae. The use of MHT alone is not recommended to confirm the presence of carbapenemases produced by Enterobacteriaceae. On the other hand Carba NP test can be used for this purpose, however molecular analysis should be considered for suspicious negative results.